Transcriptional alterations may play a role in leukemic escape and relapse following allo-HSCT

Allogeneic hematopoietic stem cell transplantation (allo-HCST) is an effective therapy option for the treatment of acute myeloid leukemia (AML), comprising high-dose chemotherapy and cancer immunotherapy, where donor immune cells eradicate leukemic cells, a phenomenon called graft-versus-leukemia effect. However, the risk of relapse remains following allo-HSCT, representing the leading cause of death post allo-HSCT. Genomic loss of human leukocyte antigen (HLA) has previously been shown to play a role in leukemia escape, and ultimately relapse. Christina Toffalori et al.¹ conducted a study to elucidate the possible contributors to allo-HSCT relapse based on genomic profiling of AML blast cells obtained from patients with AML without genomic loss of HLA at subsequent time points throughout the course of disease. The results of this analysis have been presented by Christina Toffalori during the 46th Annual Meeting of the European Society for Blood and Marrow Transplantation,² and we are pleased to summarize the key points.

Methods

With the aim of comparatively monitoring the genomic and transcriptional changes by single-nucleotide polymorphism arrays and genome-wide microarrays, two independent cohorts of adult patients with AML who underwent allo-HSCT were identified: discovery cohort (n = 40) and validation cohort (n = 36). The samples were collected at diagnosis, at the time of relapse following chemotherapy, and at the time of allo-HSCT to compare the leukemic blast cell profile at different time points^1,2.

The median age at the time of transplantation was 47 years (29–74) and 55 years (21–76) in discovery and validation cohorts, respectively. The majority of patients across both cohorts had peripheral blood as the graft source for stem cell transplantation (82.5% and 80.6%, respectively). Patient characteristics are presented in Table 1.

Table 1. Patient characteristics among two cohorts²

Results

The comparison of single-nucleotide polymorphism arrays and genomic expression profiling between the leukemic cells obtained at the time of diagnosis and at relapse after allo-HSCT in the discovery cohort demonstrated that 62% of deregulated biologic processes were immune related.

In the same samples, semantic similarity analysis was done to identify two main clusters of genes related to (1) T cell costimulation and (2) antigen processing and presentation through HLA class II molecules.

T cell costimulation

For T cell costimulation cluster, paired leukemia samples (n = 33) were assessed by flow cytometry; increased levels of expression were observed in inhibitory molecules at the time of relapse after allo-HSCT. The most significant results were increases in PD-L1, B7-H3, and PVRL2 expression and decreased expression of the T cell activation molecule, CD11A. On the same samples, CD3⁺ T cell expression of cognate receptors were studied. This found that PD-1 levels were significantly higher in patients with AML than controls, and there was a loss of CD28 expression. This indicates there was a significant change in the costimulatory link between donor T cells and leukemic cells between the two time points, e.g., CD28:CD80, ICAM1:CD11A.

HLA class II molecules

The results of mRNA expression in antigen presentation through HLA class II molecules demonstrated a consistent downregulation in the majority of HLA class II molecules and their regulator, CIITA, after allo-HSCT. The analysis of surface protein expression supported the loss of HLA-DR and HLA-DP in leukemic cells.

Gene cluster analysis of discovery and validation cohorts confirmed upregulation of coinhibitory ligands, such as PD-L1, and downregulation of HLA class II surface expression in leukemic blast cells between the time of diagnosis and relapse after transplant. This study further demonstrated that the mechanisms of downregulation of HLA class II and upregulation of coinhibitory ligands did not overlap and were irrespective of type of transplantation, i.e., donor–recipient match or mismatch. Overall, these two mechanisms of deregulation were considered responsible for up to 50% of relapses occurring following transplantation.

Therapeutic implications

Both mechanisms were associated with genes regulated by interferon (IFN). The upregulation of inhibitory ligands was aggravated by inflammation and could be blocked via immune checkpoint molecules. In an in vitro study, donor T cells responded better to leukemic cells in the presence of an anti-PDL1 monoclonal antibody. In addition, downregulation of HLA class II molecules on leukemia cells was recovered with inflammation and was restored with IFN-ɣ exposure. In co-culture, donor T cells obtained from a patient in relapse involving the depletion of HLA class II expression were found to respond to leukemia cells at the time of diagnosis but lacked response at the time of relapse. This could be recovered when leukemia cells were exposed to IFN-ɣ.

Conclusion

This study has demonstrated the possible impact of transcriptional changes on the leukemia escape and relapse following allo-HSCT along with genomic alterations. Two mechanisms that show opposite response to inflammation, i.e., the upregulation of inhibitory ligands and the downregulation of HLA class II expression, may be associated with the ability of leukemia cells to escape from donor T cells and terminate the immune response. The deregulation of allorecognition related to T cells may be a unique characteristic of post-transplantation relapse. IFN-ɣ release via induction of mild graft-versus-host disease or by blocking an immune checkpoint may be considered suitable approaches to manage these mechanisms.